krn-7000 and Carcinoma--Lewis-Lung

Attempts to harness mouse type I NKT cells in different therapeutic settings including cancer, infection, and autoimmunity have proven fruitful using the CD1d-binding glycolipid α-galactosylceramide (α-GalCer). In these different models, the effects of α-GalCer mainly relied on the establishment of a type I NKT cell-dependent immune cascade involving dendritic cell, NK cell, B cell, or conventional CD4(+) and CD8(+) T cell activation/regulation as well as immunomodulatory cytokine production. In this study, we showed that γδ T cells, another population of innate-like T lymphocytes, displayed a phenotype of activated cells (cytokine production and cytotoxic properties) and were required to achieve an optimal α-GalCer-induced immune response. Using gene-targeted mice and recombinant cytokines, a critical need for IL-12 and IL-18 has been shown in the α-GalCer-induced IFN-γ production by γδ T cells. Moreover, this cytokine production occurred downstream of type I NKT cell response, suggesting their bystander effect on γδ T cells. In line with this, γδ T cells failed to directly recognize the CD1d/α-GalCer complex. We also provided evidence that γδ T cells increase their cytotoxic properties after α-GalCer injection, resulting in an increase in killing of tumor cell targets. Moreover, using cancer models, we demonstrated that γδ T cells were required for an optimal α-GalCer-mediated anti-tumor activity. Finally, we reported that immunization of wild-type mice with α-GalCer enhanced the adaptive immune response elicited by OVA, and this effect was strongly mediated by γδ T cells. We conclude that γδ T cells amplify the innate and acquired response to α-GalCer, with possibly important outcomes for the therapeutic effects of this compound.

Topics: Adaptive Immunity; Animals; Antigens, CD1d; Carcinoma, Lewis Lung; Cytotoxicity, Immunologic; Galactosylceramides; Immunity, Innate; Interleukin-12; Interleukin-18; Killer Cells, Natural; Liver; Lung Neoplasms; Lymphocyte Activation; Lymphoma, B-Cell; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Mice, Knockout; Protein Binding; Receptors, Antigen, T-Cell, gamma-delta; Spleen; T-Lymphocytes

Alpha-Galactosylceramide (alpha-GalCer) is a potent CD1d ligand that activates natural killer like T-cells (NKT), leading to the production of helper T (Th) 1 and Th2 cytokines that mediate various immunemodulatory and antitumor effects. Here, we determined whether the administration of adenovirus-vector-encoding mouse interleukin-2 (AdmIL-2) can augment the antitumor effect of alpha-GalCer on subcutaneous and metastatic tumors in mice. Mice were intraperitoneally injected with alpha-GalCer on days 7, 10 and 13 after tumor inoculation, with or without intratumoral injection of AdmIL-2 on day 7. alpha-GalCer treatment increased the serum levels of interferon-gamma, while intratumoral injection of AdmIL-2 elevated serum IL-2 levels. A combination of alpha-GalCer and AdmIL-2 (alpha-GalCer/AdmIL-2) inhibited the in vivo tumor growth and improved the survival of tumor-bearing mice, as compared to the use of a single agent. Experiments on spontaneous metastasis models revealed that alpha-GalCer/AdmIL-2 reduced lung metastasis and prolonged survival, as compared to control groups. In addition, the splenic and liver mononuclear cells from mice treated with alpha-GalCer/AdmIL-2 showed enhanced cytolytic activity against NK-sensitive YAC-1 and NK-resistant 3LL tumors. Moreover, alpha-GalCer/AdmIL-2 treatment expanded the absolute numbers of lung and liver NK, NKT and T-cells as well as the TNF-related apoptosis-inducing ligand (TRAIL) expression of these cells. This study shows the efficacy of alpha-GalCer/AdmIL-2 immunomodulatory therapy, and provides a cellular mechanism on how it exerts the antitumor effects.

Topics: Adenoviridae; Animals; Carcinoma, Lewis Lung; Cell Line, Tumor; Cell Proliferation; Cytotoxicity Tests, Immunologic; Cytotoxicity, Immunologic; Female; Flow Cytometry; Galactosylceramides; Gene Transfer Techniques; Injections, Intraperitoneal; Interferon-gamma; Interleukin-2; Killer Cells, Natural; Mice; Mice, Inbred C57BL; Natural Killer T-Cells; Neoplasm Metastasis; Survival Analysis; T-Lymphocytes; TNF-Related Apoptosis-Inducing Ligand; Tumor Burden

CD1d-restricted natural killer T (NKT) cells are a potential therapeutic target for cancer, for which several clinical trials have already been reported. NKT cells are specifically activated by a synthetic glycolipid, alpha-galactosylceramide (alpha-GalCer). However, it is known that, in human cancer patients, NKT cells express a degree of hyporesponsiveness to alpha-GalCer. In this study, we have examined the mechanism by which hyporesponsiveness to alpha-GalCer can be induced. In cancer-bearing mice, alpha-GalCer-induced NKT cell expansion, cytokine production, cytotoxicity, and antimetastatic effect in vivo were all significantly impaired. In fact, alpha-GalCer could eliminate metastatic disease in naive animals but failed to protect cancer-bearing mice. CD11b(+) Gr-1(+) cells were particularly increased in cancer-bearing mice and were necessary and sufficient for the suppression of the alpha-GalCer response in a nitric oxide-mediated fashion. Administration of a retinoic acid to cancer-bearing mice reduced the population of CD11b(+) Gr-1(+) cells and effectively restored alpha-GalCer-induced protection. These results show a novel feature of NKT cell function in cancer. Furthermore, our data suggest a new strategy to enhance NKT cell-mediated anticancer immune responses by suppressing CD11b(+) Gr-1(+) cell functions.

Topics: Animals; Carcinoma, Lewis Lung; CD11b Antigen; Cell Line, Tumor; Enzyme-Linked Immunosorbent Assay; Galactosylceramides; Interferon-gamma; Interleukin-4; Killer Cells, Natural; Male; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Neoplasms, Experimental; Nitric Oxide; Receptors, Chemokine; Spleen; T-Lymphocytes; Time Factors; Tretinoin